1. Field of Invention
This invention relates to the area of immunology and virology, and specifically relates to compositions obtainable from mammalian thymus cells which are useful as diagnostics, vaccines, and therapeutics for human immunodeficiency virus (HIV) infection and related diseases such as acquired immunodeficiency syndrome (AIDS) and AIDS-related complex (ARC). The diagnostic, vaccination, and therapeutic methods, and devices, using these compositions are also disclosed.
2. Related Art
Bone marrow produces cells which are destined to become immune cells. These cells become lymphocytes or phagocytes. Lymphocytes are small white blood cells that bear the major responsibility for carrying out the activities of the immune system. The two major classes of lymphocytes are B cells and T cells. B cells mature in the bone (thus the term “B cells”) marrow. T cells migrate to the thymus (thus the term “T cells”) where they multiply and mature into cells capable of immune response. Upon exiting the bone marrow and thymus, both B and T cells travel widely and continuously throughout the body.
There are two types of T cells, regulatory and cytotoxic T cells, which contribute to the immune defenses in two major ways. Chief among the T cells are “helper/inducer” cells. Identifiable by the T4 cell marker, helper T cells are essential for activating B cells and other T cells as well as natural killer cells and macrophages. Cytotoxic T cells are killer cells which, for example, directly attack and rid the body of cells that have been infected by viruses or transformed by cancer.
Important phagocytes are monocytes and macrophages. Monocytes circulate in the blood, then migrate into tissues where they develop into macrophages (“big eaters”). Macrophages are found throughout the body tissues and are versatile cells that play many roles. As scavengers, they rid the body of worn-out cells and other debris. Foremost among cells that present antigen to T cells, having first digested and processed it, macrophages play a crucial role in initiating the immune response. As secretory cells, monocytes and macrophages are vital to the regulation of immune responses. They also carry receptors for lymphokines that allow them to be “activated” to pursue microbes and tumor cells.
Acquired Immunodeficiency Syndrome (AIDS) is caused by a virus, the human immunodeficiency virus (HIV). HIV destroys helper T cells and is harbored in macrophages and monocytes. AIDS is characterized by various unusual infections and otherwise rare cancers. HIV also damages tissue of the brain and spinal cord, producing progressive dementia. Entry of HIV-1 into helper T cells involves the primary receptor CD4 and co-receptors CCR5 and CXCR4. The first step in cell entry occurs when the HIV-1 glycoprotein gp120 binds to the CD4 receptors on target cells. The next step is an interaction between the HIV-1 envelope protein and the co-receptor CCR5. Once gp120 interacts with receptor and co-receptor, the HIV-1 envelope protein gp41 undergoes a conformational change and literally brings the viral membrane into close proximity with the cell membrane. Fusion of two lipid bilayers then occurs, allowing intracellular entry of the viral contents.
When HIV infects a human patient, it incorporates itself into the deoxyribonucleic acid (DNA) of the immune cells and for a variable period of between 3 months to years, the patient may not exhibit any immunodeficiency symptoms and sometimes does not produce a detectable level of antibodies against AIDS. Since an initial HIV infection may not immediately lead to detectable clinical disease symptoms or a detectable level of antibodies, the term “HIV infection” as used herein encompasses both the infection and any disease resulting therefrom, the latter being termed “HIV-related diseases”. Examples of HIV-related diseases are AIDS and ARC. After the above incubation period, the HIV multiplies within the infected cell and eventually bursts the host cells which release the newly formed viruses. Since the host cells are destroyed in the process, the patient's immune system is impaired and the host is susceptible to opportunistic diseases that a human with intact immune system is not susceptible to. In human, generally the AIDS virus will multiply and the human will eventually die from severe immunodeficiency. Interestingly, only humans suffer from AIDS. When a non-human mammal, such as a rabbit, mouse, rat or cow, is injected with HIV, the animal may temporarily have some T cells destroyed. However, 14 to 21 days post-infection, the animal would mount an antibody attack and does not succumb to AIDS. Thus, there is no animal model for AIDS.
Currently, there is no cure or effective therapeutic for AIDS. Research in this area is underway.
There is also a continuous search for a method for accurately diagnosing AIDS at an earlier stage of the disease. Currently, the commercially available diagnostic tests are generally directed to detecting the patients antibodies against HIV. Such tests have a window of about 14 to 21 days from the time the patient is infected with AIDS to the time the patient produces antibodies against the virus. Therefore, if a patient is tested during this window of time, the tests will produce a false negative result. On the other hand, some of these tests may also give false positive results due to non-specific binding of the antibodies. Another means for detecting the viral infection is through nucleic acid hybridization.
Unless otherwise noted, the following is based on Stein, D. S, et al., . Infect. Diseases, 165: 3 52 (1992). The surrogate marker that most closely correlates with the stage of HIV infection is the CD4+, or T helper, cell count. HIV-1 envelope glycoprotein, gp120, specifically binds to the CD4 receptor that is expressed in greatest concentration in a subset of T lymphocytes and in lower amounts on monocytes and macrophages. Cells expressing CD4 receptors are termed the “helper/inducer” subset, reflecting their role as both helper cells for B cell responses for antigens expressed on cells bearing human leukocyte antigen (HLA) class II receptors and inducer cells that cause T cells to suppress immune responses. The selective loss of CD4+ cells results in numerous immune defects associated with susceptibility to the opportunistic infections that are the hallmark of AIDS.
The HIV core antigen p24 can be detected before the appearance of HIV antibodies. After the appearance of HIV antibodies by the screening enzyme-linked immunosorbent assay (ELISA), p24 antigenemia generally becomes undetectable, though it can occasionally persist and often will recur later in the disease. HIV-I titers found in plasma and peripheral blood mononuclear cell cultures also fall rapidly as specific antibodies are detectable, suggesting at least a transiently effective host immune response. Markers of immune stimulation includes β2-microglobulin.
In patients followed from the time of seroconversion, CD4+ cell decline has been correlated with progression to AIDS. Serum levels of β2-microglobulin and detection of p24 antigen in blood were also both independently correlated with rates of progression. Combined with CD4 cell counts, use of β2-microglobulin and p24 antigen increased prognostic accuracy for progression to AIDS compared with CD4+ cell count alone.
However, it was rare for seroconverters to have a consistent decline in their percentage of CD4+ cells over the next three years. In the interval between visits, stable or declining levels of CD4+ cell percentages were found in 38% of subjects, with 12% experiencing declines followed by a leveling in their rates of loss of CD4 cells. Overall, 62% experienced declines in their CD4+ cell percentage over three years of follow-up.
In a study of 306 HIV-infected seropositive homosexual men with unknown times of seroconversion, both a CD4+ cell count <500/μl and p24 antigen detection were predictive of AIDS within 30 months.
Increased CD8+ cell counts were found to be somewhat predictive of subsequent development of AIDS.
To better correlate clinical end points, such as survival and progression to AIDS, with surrogate markers of antiviral therapy effects, analysis of additional markers such as neopterin and β2-microglobulin, among others, have been combined with the CD4 cell count and p24 antigen.
In a limited study {Jacobson, M. A., BNJ, 302:73 (1991)} of patients with AIDS and ARC who tolerated an anti-AIDS drug, zidovudine, and who survived for 12 weeks, the following was found.
After controlling for three factors (age, diagnosis of AIDS at baseline, log of the baseline serum neopterin concentration), the log of the CD4+ cell count at 8-12 weeks, but not the change over time, best predicted subsequent survival. A decrease in β2-microglobulin concentration at 8-12 weeks significantly predicted survival and, combined with the log of the CD4+ cell count, provided the best predictive model. Decreases in p24 antigenemia, serum neopterin concentrations, and the Karhofsky performance status (a measure of function in routine activities) did not significantly correlate with survival on therapy.
Stein, D. S., et al., above, conclude that changes in CD4+ cell counts and other surrogate markers may be increasingly used as the sole end point for investigations of antiretroval activity, of a drug or therapy, in patients with early HIV infection.